Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells

Endosomal sorting complex required for transport (ESCRT) machinery supports the efficient budding of Marburg virus (MARV) and many other enveloped viruses. Interaction between components of the ESCRT machinery and viral proteins is predominantly mediated by short tetrapeptide motifs, known as late d...

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Main Authors:	Dolnik, Olga, Kolesnikova, Larissa, Welsch, Sonja, Strecker, Thomas, Schudt, Gordian, Becker, Stephan
Format:	Online
Language:	English
Published:	Public Library of Science 2014
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4199773/

id	pubmed-4199773
recordtype	oai_dc
spelling	pubmed-41997732014-10-21 Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells Dolnik, Olga Kolesnikova, Larissa Welsch, Sonja Strecker, Thomas Schudt, Gordian Becker, Stephan Research Article Endosomal sorting complex required for transport (ESCRT) machinery supports the efficient budding of Marburg virus (MARV) and many other enveloped viruses. Interaction between components of the ESCRT machinery and viral proteins is predominantly mediated by short tetrapeptide motifs, known as late domains. MARV contains late domain motifs in the matrix protein VP40 and in the genome-encapsidating nucleoprotein (NP). The PSAP late domain motif of NP recruits the ESCRT-I protein tumor susceptibility gene 101 (Tsg101). Here, we generated a recombinant MARV encoding NP with a mutated PSAP late domain (rMARVPSAPmut). rMARVPSAPmut was attenuated by up to one log compared with recombinant wild-type MARV (rMARVwt), formed smaller plaques and exhibited delayed virus release. Nucleocapsids in rMARVPSAPmut-infected cells were more densely packed inside viral inclusions and more abundant in the cytoplasm than in rMARVwt-infected cells. A similar phenotype was detected when MARV-infected cells were depleted of Tsg101. Live-cell imaging analyses revealed that Tsg101 accumulated in inclusions of rMARVwt-infected cells and was co-transported together with nucleocapsids. In contrast, rMARVPSAPmut nucleocapsids did not display co-localization with Tsg101, had significantly shorter transport trajectories, and migration close to the plasma membrane was severely impaired, resulting in reduced recruitment into filopodia, the major budding sites of MARV. We further show that the Tsg101 interacting protein IQGAP1, an actin cytoskeleton regulator, was recruited into inclusions and to individual nucleocapsids together with Tsg101. Moreover, IQGAP1 was detected in a contrail-like structure at the rear end of migrating nucleocapsids. Down regulation of IQGAP1 impaired release of MARV. These results indicate that the PSAP motif in NP, which enables binding to Tsg101, is important for the efficient actin-dependent transport of nucleocapsids to the sites of budding. Thus, the interaction between NP and Tsg101 supports several steps of MARV assembly before virus fission. Public Library of Science 2014-10-16 /pmc/articles/PMC4199773/ /pubmed/25330247 http://dx.doi.org/10.1371/journal.ppat.1004463 Text en © 2014 Dolnik et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Dolnik, Olga Kolesnikova, Larissa Welsch, Sonja Strecker, Thomas Schudt, Gordian Becker, Stephan
spellingShingle	Dolnik, Olga Kolesnikova, Larissa Welsch, Sonja Strecker, Thomas Schudt, Gordian Becker, Stephan Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
author_facet	Dolnik, Olga Kolesnikova, Larissa Welsch, Sonja Strecker, Thomas Schudt, Gordian Becker, Stephan
author_sort	Dolnik, Olga
title	Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
title_short	Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
title_full	Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
title_fullStr	Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
title_full_unstemmed	Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells
title_sort	interaction with tsg101 is necessary for the efficient transport and release of nucleocapsids in marburg virus-infected cells
description	Endosomal sorting complex required for transport (ESCRT) machinery supports the efficient budding of Marburg virus (MARV) and many other enveloped viruses. Interaction between components of the ESCRT machinery and viral proteins is predominantly mediated by short tetrapeptide motifs, known as late domains. MARV contains late domain motifs in the matrix protein VP40 and in the genome-encapsidating nucleoprotein (NP). The PSAP late domain motif of NP recruits the ESCRT-I protein tumor susceptibility gene 101 (Tsg101). Here, we generated a recombinant MARV encoding NP with a mutated PSAP late domain (rMARVPSAPmut). rMARVPSAPmut was attenuated by up to one log compared with recombinant wild-type MARV (rMARVwt), formed smaller plaques and exhibited delayed virus release. Nucleocapsids in rMARVPSAPmut-infected cells were more densely packed inside viral inclusions and more abundant in the cytoplasm than in rMARVwt-infected cells. A similar phenotype was detected when MARV-infected cells were depleted of Tsg101. Live-cell imaging analyses revealed that Tsg101 accumulated in inclusions of rMARVwt-infected cells and was co-transported together with nucleocapsids. In contrast, rMARVPSAPmut nucleocapsids did not display co-localization with Tsg101, had significantly shorter transport trajectories, and migration close to the plasma membrane was severely impaired, resulting in reduced recruitment into filopodia, the major budding sites of MARV. We further show that the Tsg101 interacting protein IQGAP1, an actin cytoskeleton regulator, was recruited into inclusions and to individual nucleocapsids together with Tsg101. Moreover, IQGAP1 was detected in a contrail-like structure at the rear end of migrating nucleocapsids. Down regulation of IQGAP1 impaired release of MARV. These results indicate that the PSAP motif in NP, which enables binding to Tsg101, is important for the efficient actin-dependent transport of nucleocapsids to the sites of budding. Thus, the interaction between NP and Tsg101 supports several steps of MARV assembly before virus fission.
publisher	Public Library of Science
publishDate	2014
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4199773/
_version_	1613145733131665408

Interaction with Tsg101 Is Necessary for the Efficient Transport and Release of Nucleocapsids in Marburg Virus-Infected Cells

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